FUNCTIONAL CONSEQUENCES IN YEAST OF SINGLE-RESIDUE ALTERATIONS IN A CONSENSUS CALMODULIN

A synthetic gene encoding a 'consensus' calmodulin (synCaM) was able t o substitute for the Saccharomyces cerevisiae calmodulin gene (CMD1), even though synCaM is only 60% identical in primary amino acid sequenc e to yeast Cmd1. Twelve different synCaM mutants were also expressed i n yeast. Seven of the 12 mutant synCaMs supported germination and grow th of Cmd1-deficient spores. Five of the 12 mutant synCaMs were incapa ble of supporting germination of Cmd1-deficient spores and, of these, four were also incapable of supporting vegetative growth of Cmd1-defic ient haploid cells. The five nonfunctional synCaM mutants were express ed at levels equivalent to, or higher than, the seven synCaM mutants t hat were able to substitute for Cmd1; thus, the inability to function was not simply due to inadequate expression or rapid degradation. All nonfunctional synCaM mutants shared a single charge reversal mutation in the central helix (E84K), which was found to be sufficient to confe r the lethal phenotype. The ability of another mutant synCaM (S101F) t o support growth of Cmd1-deficient cells was dependent on cell ploidy. Another mutant (K115Y) supported spore germination and vegetative gro wth, but not meiosis and sporulation. The terminal phenotype of cells lacking a functional calmodulin included a dramatic accumulation of po lymerized microtubules.